In a number of aspects the work proposed here will be a continuation and extension of our work originally initiated 15 to 20 years ago, and supported by GM-12394 since 1964 under the title given above. This title does indeed state our principal area of past contributions and the main thrust of our proposed work exactly. With the idea in mind that the one technique (viz. EPR spectroscopy) which permits direct and quantitative determination of unpaired electrons should be able to provide new and decisive insights in the area of biological electron transfer, we have systematically adopted and further developed ways and means to apply this and auxiliary techniques to biological problems. This will continue to be one of our main aims. Our emphasis will be less on highly purified systems than in the past, and more on complex systems such as mitochondria of various species, membranous subfractions, and, in some instances, whole tissue. Unknown components of these systems that are capable of accepting electrons will be characterized by detailed study after isolation, if this is feasible, by comparison to optical features, by oxidoreductive titrations and rapid kinetic studies and, possibly, by the use of specific inhibitors. As our knowledge of such systems developed in the past, an increasing number of initially unexplained features in EPR spectra could be explained by interactions of paramagnetic components. On the basis of experience gathered to date we will give such features more attention than in the past. Our main aim in the development of apparatus suitable for experimentation in thus far unexploited areas will be to develop a reliable and practical system for preparing samples for EPR spectroscopy by rapid freezing in aqueous-organic solvent mixtures at subzero temperatures. This technique should bring within the range of EPR spectroscopic examination a large number of transient phenomena and intermediates which have thus far been inaccessible. We expect that there will be demand for our collaboration in this approach by a considerable number of colleagues.